This invention relates to methods for determining an analyte in a blood sample based on latex agglutination immunoassay. More particularly, the invention concerns a method for overcoming nonspecific agglutination from hemoglobin present in a test sample. The invention is particularly directed to the determination of glycated hemoglobin, e.g., hemoglobin Alc, in samples of whole blood.
Latex agglutination immunoassay is based on the formation of detectable agglutination by binding between a multivalent latex antibody reagent and a corresponding multivalent form of the antigen or hapten. Where an analyte of interest is itself multivalent relative to the latex antibody reagent, a direct assay can be conducted. However, where the analyte of interest is not multivalent, such as in the case of a low molecular weight hapten, a competitive assay is performed in order to quantitate such haptenic analyte. An agglutinator reagent is added to the system comprised of a plurality of epitopic binding sites for the anti-analyte antibody reagent. Binding between the agglutinator reagent and the latex antibody reagent results in formation of detectable agglutination. With increasing amounts of analyte, which competes with the agglutination reagent for binding to the latex antibody reagent, the amount of resulting agglutination is reduced.
The prior art has applied the latex agglutination immunoassay method principally to serum or plasma samples in order to determine analytes of interest in blood. It has been found that the presence of whole blood components can give rise to nonspecific agglutination of the latex reagent. Therefore, in situations where it is desirable or necessary to assay whole blood, the precision of the assay result is subject to possible large errors. This particularly applies to the determination of blood components associated with erythrocytes such as hemoglobin derivatives of diagnostic importance, e.g., the glycated hemoglobins.